Development of a technology to certify engineered DNA molecules

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PROJECT SUMMARY The open availability of authenticated, well-documented research materials is essential for scientific progress. Plasmids have become essential research tools to address almost any question in biology, and together with other forms of engineered DNA molecules, are essential for clinical research applications including gene therapy, vaccine development, and the production of recombinant drugs. Currently, the two common links between a plasmid and its documentation are the plasmid names and the plasmid sequence. Despite the central role that plasmids play in biomedical research and development, there is no guaranteed way to connect a physical plasmid in a tube to its documentation. A pipetting error, a labelling error, a spontaneous mutation, or an undocumented modification of the plasmid are some of the events that could result in a tube containing a different plasmid than what is indicated on the label. In addition, there is no standardized, secure approach to documenting the sequence, function, and lineage of a plasmid. As a result, there are widespread discrepancies between the physical sequences of the plasmids in circulation in the life science community and their supposed reference sequence. This situation creates reproducibility issues, slows down R&D efforts and raises significant security and safety issues for biotechnology applications. We are proposing to develop a new digital certificate technology, enabled by a web-based resource called MyPlasmid.org, that will provide a robust, physical link between engineered DNA molecules, their electronic documentation, and their authors. This technology will produce unique DNA sequences generated by cryptographic algorithms that can be inserted into an engineered DNA molecule. MyPlasmid.org will allow users to document their genetic designs by aggregating the documentation of individual genetic elements as well as combinations of elements. In addition, it will link the computer records of the engineered DNA sequence directly to the molecule itself and provide a method to retrieve documentation without a priori knowledge of the plasmid's identity. Short unique DNA sequences called certificates will be inserted between the functional blocks of engineered sequences. Unlike DNA barcodes, certificates will be computed by cryptographic algorithms using the DNA sequence itself and the author's identity as input so that users of engineered DNA molecules can verify the origin and integrity of certified DNA molecules. The technology described in this proposal is expected to foster a transition similar what has been observed in the semi-conductor industry where different stakeholders invest in the development of circuits that can be easily combined in larger designs that can then be manufactured by foundries not involved in the chip design. By ensuring that the sequence, origin, function, and lineage of engineered DNA molecules is accurately tracked and conveyed to all users, the proposed technology will improve the reproducibility, utility, and potential application of engineered DNA molecules in the life sciences.